琥珀酸脱氢酶或琥珀酰辅酶A合成酶缺失促进大肠杆菌积累5-氨基乙酰丙酸

Deficiency of succinic dehydrogenase or succinyl-CoA synthetase enhances the production of 5-aminolevulinic acid in recombinant Escherichia coli

摘要:

5-氨基乙酰丙酸 (ALA) 是生物体内四吡咯类化合物的合成前体，在农业及医药领域应用广泛，是极具开发价值的高附加值生物基化学品。目前利用外源C4途径的重组大肠杆菌发酵生产ALA的研究主要利用LB培养基并添加葡萄糖和琥珀酸、甘氨酸等合成前体，成本较高。琥珀酸在C4途径中以琥珀酰辅酶A的形式直接参与ALA的合成。文中在以葡萄糖为主要碳源的无机盐培养基中研究了琥珀酰辅酶A下游代谢途径琥珀酸脱氢酶编码基因sdhAB和琥珀酰辅酶A合成酶编码基因sucCD缺失对ALA积累的影响。与仅表达异源ALA合成酶的对照菌株相比，sdhAB和sucCD缺失菌株ALA的产量分别提高了25.59%和12.40%，且ALA的积累不依赖于琥珀酸的添加和LB培养基的使用，从而大幅降低了生产成本，显示出良好的工业应用前景。

关键词:5-氨基乙酰丙酸，琥珀酸脱氢酶，琥珀酰辅酶A合成酶，大肠杆菌，基因敲除

Abstract:

5-aminolevulinic acid (ALA), a precursor for biosynthesis of pyrrole compounds in living organisms, has been widely used in agriculture and medical photodynamics therapy and is regarded as a promising value-added bio-based chemical. In the previous investigations on ALA production with recombinant Escherichia coli expressing heterogenous C4 pathway gene, LB media supplemented with glucose and ALA precursors succinate and glycine is widely used, leading to high production cost. Succinate participates in ALA biosynthesis in a form of succinyl-CoA. In this study, genes involved in succinyl-CoA consumption, sdhAB (encoding succinic dehydrogenase) or sucCD (encoding succinyl-CoA synthetase) of E. coli MG1655 was knocked out and tested for ALA accumulation. In comparison with the recombinant E. coli strain expressing heterogenous ALA synthetase, the sdhAB- or sucCD-deficient strain accumulate 25.59% and 12.40%, respectively, more ALA in a 5 L fermentor using a defined synthetic medium with glucose as main carbon source and without supplementation of succinate, providing a novel cost-effective approach for industrial production of ALA.

Key words:5-aminolevulinic acid, succinic dehydrogenase, succinyl-CoA synthetase, Escherichia coli, gene knock out